Ghosts of symbionts past: The hidden history of the dynamic association between filarial nematodes and their Wolbachia endosymbionts: Supplementary Data files

Supplementary data, Figures and Tables for manuscript "Ghosts of symbionts past: The hidden history of the dynamic association between filarial nematodes and their Wolbachia endosymbionts"

Emmelien Vancaester, Guy R. Oldrieve, Alex Reid, Georgios Koutsovoulos, Dominik R. Laetsch, Benjamin L. Makepeace, Vincent Tanya, Sven Poppert, Jürgen Krücken, Adrian Wolstenholme, Mark Blaxter

Many, but not all, parasitic filarial nematodes (Onchocercidae) carry intracellular, maternally-transmitted Wolbachia symbionts, and these alphaproteobacteria are targets for anti-filarial chemotherapeutic interventions for human disease. The symbionts of Onchocercidae derive from four of the major supergroups (C, D, F and J) defined within the genus Wolbachia. Using twenty-two whole genome sequences of filarial nematodes and genome sequences of their Wolbachia partners, we have explored the evolutionary history of the nematode-Wolbachia symbiosis. We screened the nuclear genome sequences of all the nematodes for nuclear Wolbachia transfers (NUWTs), fragments of the Wolbachia genome that have been integrated into the nuclear genome. 

Six species have no current Wolbachia infection. Setaria labiatopapillosa had no validated NUWTs and we interpret this to mean that this species was never infected with Wolbachia. In the other five species (Acanthocheilonema viteae, Cercopithifilaria (Ce.) johnstoni, Elaeophora elaphi, Loa (Lo.) loa and Onchocerca flexuosa) we found NUWTs, implying they have previously had and have now lost Wolbachia infections. For each NUWT locus, we identified the supergroup membership of the Wolbachia from which it originated, and found that the five Wolbachia-free species carried NUWTs derived from multiple supergroups, including a high shared rate of sequences derived from supergroup C. In Dirofilaria repens we identified a sample that carried two Wolbachia symbionts, one from supergroup C and one from supergroup F. In Dirofilaria immitis where live infection with a supergroup C Wolbachia is found, we identified NUWTs derived from an F Wolbachia, confirming that the F association predated the divergence of these Dirofilaria species. The supergroup D lineage of Wolbachia, as present in the human parasites Wuchereria bancrofti and Brugia malayi, derives from a replacement event. Madathamugadia (Md.) hiepei shows signs of multiple recent repeated endosymbiont replacement.

From these data we infer that the history of Wolbachia in onchocercid nematodes includes not only cospeciation (as present in the Onchocerca-Dirofilaria group in association with supergroup C Wolbachia) and loss (in the five Wolbachia-free species), but also frequent symbiont replacement and dual infection. This dynamic pattern is challenging to models that assume host-symbiont mutualism.

The zip archive contains:

Scripts

Python scripts written for data processing.

FetchRegionSeq.py.gz

Reduce_alignment.py.gz

Reroot_rename_tree.py.gz

SelectMatches.py.gz

Data

Directories containing intermediate data files generated for the analyses presented.

1_WolbachiaAnnotation - The prokka annotation of each Wolbachia genome analysed

2_WolbachiaOrthoFinder - The orthoFinder results for the clustering of the proteomes of the 165 selected Wolbachia proteomes

3_WolbachiaPhylogeny - The phylogeny of Wolbachia inferred using Astral 

4_WolbachiaOrthoAlignments - Protein alignments of orthogroups from the orthoFinder analysis of 165 selected Wolbachia peoteomes

5_WolbachiaHMM - nucleotide hidden Markov models generated from the nucleotide alignments of orthogroups from the orthoFinder analysis of 165 selected Wolbachia proteomes

6_NUWTtrees - Phylogenies inferred using Astral of the Wolbachia orthogroups for which NUWT sequences were identified in the filarial nematode genomes. The NUWT sequences were profile aligned to the orthogroup nucleotide alignments.

7_NUWTmatches - File giving the nucleotide coordinates and scores of each orthogroup HMM in the filarial genomes. Tab delimited: columns are: Orthogroup number, Species abbreviation, Supergroup of origin of NUWT, Location of NUWT (Species, Contig/scaffold, base coordinates of match)

8_OnchocercidPhylogeny - Data supporting the onchocercid nematode phylogeny, including files of aligned orthogroups used in coalescent analyses in ASTRAL and the supermatrix of these alignments used for iQtree analysis.

9_New_Nematode_and_Wolbachia_Genomes - Fasta format files of the genome sequences of Setaria labiatopapillosa (INSDC accession GCA_966190365) and the Wolbachia genomes from Dirofilaria repens wDreP_C (ERZ27399396) and wDrepF (ERZ27399397).

Supplementary Tables

For completeness, the supplementary tables are replicated here.

Table S1: Nematode genome data metrics

Table S2: Wolbachia genome data

Table S3: Dereplicated Wolbachia genomes

Table S4: Contigs removed from filarial nematode genomes as likely contaminants.

Table S5: Numbers and classification of NUWTs detected in each filarial nematode species genome

Table S6: Span of NUWTs detected in each filarial nematode species’ genome.

Table S7: Average length of NUWTs.

Table S8: NUWTs from different Wolbachia supergroups.

Supplementary Figures

in a separate file

Figure S1: BlobTools plot of initial Dirofilaria repens assembly
Figure S2: Genome phylogeny of Wolbachia
Figure S3: Wolbachia proteome clustering
Figure S4: Phylogenetic tree of orthologous family OG0000236 and NUWTs
Figure S5: Phylogenetic tree of orthologous family OG0000277 and NUWTs
Figure S6: Phylogenetic tree of orthologous family OG0000301 and NUWTs
Figure S7: Phylogenetic tree of orthologous family OG0000453 and NUWTs
Figure S8: Phylogenetic tree of orthologous family OG0000206 and NUWTs